We have employed two approaches to study glucocorticoid resistance in human leukemic cells in culture. We have: (1)\constructed somatic hybrids between resistant and sensitive cells and studied the dominant or recessive behavior of the resistant phenotypes; and (2)\developed techniques for physical identification of the human IM-9 glucocorticoid receptor. Using polyethylene glycolmediated cell fusion, somatic hybrids were constructed between steroid-sensitive and either receptorless or activation labile steroid-resistant mutants. When receptor content in the hybrids was measured, it was found that hybrids contained essentially the arithmetic sum of the receptors found in each of the parents. Thus, there do not appear to be any transdominant factors in the resistant mutants which interfere with or alter normal receptor function. All of our results are consistant with the interpretation that resistance is the result of mutation within the gene(s) encoding the glucocorticoid receptor, and that the two phenotypes are manifestations of the two different mutational events at the same locus. Results of studies on IM-9 receptors suggest that either the entire receptor structure is required for antibody recognition or that the steroid binding site and the antibody recognition site are on opposite ends of the receptor molecule. Some of our mutants are defective in activation of the steroid receptor complex. We have begun a characterization of the unactivated and activated forms of the IM-9 steroid receptor complex. These data suggest that the difference between the two forms may involve conformational changes or subunit disassembly. The nature of this difference is still being investigated. Further objectives are: (1)\to define, by protein-protein crosslinking and in vitro proteolysis, the nature of the 7.2 to 5.8 nm transition associated with activation of the steroid-receptor complex; (2)\to complete biochemical characterization of the somatic hybrids; (3)\to map the human glucocorticoid receptor gene to a specific human chromosome; and (4)\to extend our studies on IM-9 glucocorticoid receptors to CEM wild-type and resistant cells using immunochemical and biochemical approaches.